The objective of this grant application is to define how Haemophilus influenzae bacteria carried within the lung adapt to evade host innate responses. The long-term objective is to understand how mimicry of host structures contributes to bacterial persistence at mucosal surfaces. H. influenzae is carried in the human airways and causes opportunistic infections such as bronchitis, sinusitis, and otitis media that are major public health problems. Innate responses to H. influenzae are largely mediated by the Toll-like receptor 4 pathway. Toll-like receptor 4 recognizes a structural pattern found in endotoxins of most gram-negatives, including H. influenzae. H. influenzae has structurally diverse lipooligosaccharide endotoxins that share certain structural modifications, including the addition of host structures like sialic acid and phosphorylcholine. Data included in this application show that the addition of phosphorylcholine to these endotoxins decreases innate responses and affinity to lipopolysaccharide binding protein, and promotes bacterial persistence in the lung. Therefore, the hypotheses of this project are: 1) innate defenses select for bacterial variants that elicit reduced innate responses, 2) these variants are more persistent, and 3) phosphorylcholine addition to LOS is such an adaptation. In order to address these hypotheses, we will complete the following Specific Aims: Specific Aim 1. To define how chronic carriage in vivo influences the composition and bioactivity of H. influenzae lipooligosaccharides. Specific Aim 2: To define the mechanism(s) whereby phosphorylcholine reduces host innate responses to H. influenzae lipooligosaccharides. With ever-increasing levels of antibiotic resistance among most bacteria, our ability to treat bacterial disease with conventional means is gradually diminishing. If we are to design new ways to prevent or treat bacterial disease, a key step will be to define how these bacteria resist clearance by host defenses in vivo. The completion of these Specific Aims will provide insights into one way that a host-adapted commensal and important opportunistic pathogen can blunt innate defenses and resist clearance. Therefore, this project offers an important opportunity to develop new ways to combat chronic airway infections that are a major cause of morbidity and mortality worldwide.